Bacteriocins are bacterial proteins produced to prevent the growth of competing microorganisms in a particular biological niche. A preparation of bacteriocin from a particular strain of E. coli (HSC.sub.10) has long been shown to have anti-neoplastic activity against a variety of human tumour cell lines in vitro (1,2). This preparation, previously referred to as PPB (partially purified bacteriocin (2)) or ACP (anti-cancer proteins (2)) was also effective in a murine tumour model of preventing metastases to the lung (2).
Verotoxins, also known as SHIGA-like toxins, comprise a family known as Verotoxin 1, Verotoxin 2, Verotoxin 2c and Verotoxin 2e of subunit toxins elaborated by some strains of E. coli (3). These toxins are involved in the etiology of the hemolytic uremic syndrome (3,4) and haemorrhagic colitis (5). Cell cytotoxicity is mediated via the binding of the B subunit of the holotoxin to the receptor glycolipid, globotriaosylceramide, in sensitive cells (6).
The verotoxin family of E coli elaborated toxins bind to the globo series glycolipid globotriaosylceramide and require terminal gal .alpha.-1-4 gal residue for binding. In addition, VT2e, the pig edema disease toxin, recognizes globotetraosylceramide (Gb.sub.4) containing an additional .beta. 1-3 linked galNac residue. These glycolipids are the functional receptors for these toxins since incorporation of the glycolipid into receptor negative cells renders the recipient cells sensitive to cytotoxicity. The toxins inhibit protein synthesis via the A subunit--an N-glycanase which removes a specific adenine base in the 28S RNA of the 60S RNA ribosomal subunit. However, the specific cytotoxicity and specific activity is a function of the B subunit. In an in vitro translation system, the verotoxin A subunit is the most potent inhibitor of protein synthesis yet described, being effective at a concentration of about 8 pM. In the rabbit model of verocytotoxemia, pathology and toxin targeting is restricted to tissues which contain the glycolipid receptor and these comprise endothelial cells of a subset of the blood vasculature. Verotoxins have been strongly implicated as the etiological agents for hemolytic uremic syndrome and haemorrhagic colitis, microangiopathies of the glomerular or gastrointestinal capillaries respectively. Human umbilical vein endothelial cells (HUVEC) are sensitive to verotoxin but this sensitivity is variable according to cell line. Human adult renal endothelial cells are exquisitely sensitive to verotoxin in vitro and express a correspondingly high level of Gb.sub.3. However, HUS is primarily a disease of children under three and the elderly, following gastrointestinal VTEC infection. It has been shown that receptors for verotoxin are present in the glomeruli of infants under this age but are not expressed in the glomeruli of normal adults. HUVEC can be sensitized to the effect of verotoxin by pretreatment by tumour necrosis factor which results in a specific elevation of Gb.sub.3 synthesis (7,8). Human renal endothelial cells on the other hand, although they express high levels of Gb.sub.3 in culture, cannot be stimulated to increase Gb.sub.3 synthesis (8). It has been suggested that the transition from renal tissue to primary endothelial cell culture in vitro results in the maximum stimulation of Gb.sub.3 synthesis from a zero background (9). We therefore suspect that HUS in the elderly is the result of verotoxemia and a concomitant stimulation of renal endothelial cell Gb.sub.3 synthesis by some other factor, eg. LPS stimulation of serum .alpha. TNF. Thus under these conditions, the majority of individuals (excepting the very young) would not be liable to VT induced renal pathology following systemic verotoxemia.
It has also been shown that the verotoxin targets a sub-population of human B cells in vitro (10). These Gb.sub.3 containing B cells are found within the germinal centres of lymph nodes (11). It has been proposed that Gb.sub.3 may be involved in a germinal centre homing by CD19 positive B cells (12) and that Gb.sub.3 may be involved in the mechanisms of antigen presentation (13).
Elevated levels of Gb.sub.3 have been associated with several other human tumours (14-16), but ovarian tumours have not been previously investigated. Gb.sub.3 is the p.sup.k blood group antigen (17). Tissue surveys using anti-p.sup.k antisera have shown that human ovaries do not express this glycolipid (18, 19). Sensitivity to VT1 cytotoxicity in vitro has been shown to be a function of cell growth, the stationary phase cells being refractile to cytotoxicity (20). The sequence homology between the receptor binding B subunit and the human .alpha.2-interferon receptor and the B cell marker CD19 suggests that expression of Gb.sub.3 is involved in the mechanism of .alpha.2-interferon and CD19 signal transduction (12). On surface ligation, Gb.sub.3 has been shown to undergo a retrograde intracellular transport via the rough endoplasmic reticulum to the nuclear membrane (21).
The astrocytoma is the most common primary human brain tumour. The majority of astrocytomas are malignant neoplasms which infiltrate diffusely into regions of normal brain. Despite the advent of promising adjuvant therapies and drugs which have impacted positively on patient survival in other tumor types in recent times, no such promising therapy has yet been found for the patient with a malignant astrocytoma. The median survival for patients with glioblastoma multiforme, the most malignant form of astrocytoma, is approximately 12 months and accordingly, it is imperative that new therapeutic treatments for malignant astrocytomas be found.
VTs consist of a 30 kDa enzymatic A subunit which is capable of inhibiting protein synthesis. The A subunit is noncovalently associated with a pentameric 7 kDa B subunit array which binds to Gb.sub.3.
In addition to the cytotoxic effects of VTs on a wide range of cells by the A subunit inhibition of protein synthesis, recent evidence suggests that VT1. and the receptor binding B subunit alone, also induce morphological changes and DNA fragmentation characteristic of apoptosis in Gb.sub.3 -positive cells (22, 23).